THE NONCLASSICAL major histocompatibility com-plex molecule, HLA-G, is mainly expressed on cytotrophoblast cells during pregnancy, 1 where it is likely involved in the protection of the semiallogeneic fetus from maternal rejection. 2 The HLA-G gene contains eight exons that encode the peptide signal (exon 1), three extracellular domains (exons 2, 3, and 4), a transmembrane region (exon 5), and a reduced cytoplasmic tail due to the presence of a premature stop codon in exon 6. 3 The primary transcript of the HLA-G gene is alternatively spliced, resulting in at least six different HLA-G mRNAs that potentially encode four membrane-bound (HLA-G1,-G2,-G3, and-G4) and two soluble (HLA-G5 and-G6) protein isoforms. 4–6 While the full-length HLA-G1, which presents similar classical HLA class I structure, was widely investigated over the past few years at both biochemical and functional levels, 7–11 HLAG2,-G3, and-G4 remain to be well defined. Previous experiments have demonstrated that HLA-G1 transfected into HLA class I-negative cells lines, such as LCL 721.22112–15 and K562, 16–17 reaches the cell surface and is responsible for NK cytolysis inhibition. This inhibition may occur through interaction with at least three inhibitory receptors, namely ILT-2, 12, 13 p49, 15, 18 and KIR2DL4. 14 However, besides the HLA-A and-B-negative placental tissue, HLA-G expression was also reported in HLA class I-positive cells such as chorionic fetal vessels, 19 thymic epithelial cells, 20 and cytokine activated monocytes, 21, 22 as well as in some melanoma biopsies. 22, 23 In this latter situation, the expression of HLA-G2,-G3, and-G4 isoforms was strongly suggested. 24 Accordingly, to characterize these isoforms, the cDNA encoding each HLA-G isoform was transfected into an HLA-class I-positive cell line, allowing us to mimick the situation in which HLA-G is coexpressed with HLA class I molecules. Each isoform was characterized at both transcriptional and protein levels. Results showed that (1) HLA-G1,-G2,-G3, and-G4 are transcribed in the corresponding transfectant;(2) HLA-G1,-G2,-G3, and-G4 are translated in protein that could be detected by using the 4H84 monoclonal antibody (mAb), which reacts with all denatured isoforms, 20 and (3) only the HLA-G1 protein could be cell-surface detected by the use of the specific anti-HLA-G1 mAb currently available, namely 87G. 7